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Subjects

Abstract

An intronic GGGGCC repeat expansion in C9ORF72 is the most common cause of amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD), but the pathogenic mechanism of this repeat remains unclear. Using human induced motor neurons (iMNs), we found that repeat-expanded C9ORF72 was haploinsufficient in ALS. We found that C9ORF72 interacted with endosomes and was required for normal vesicle trafficking and lysosomal biogenesis in motor neurons. Repeat expansion reduced C9ORF72 expression, triggering neurodegeneration through two mechanisms: accumulation of glutamate receptors, leading to excitotoxicity, and impaired clearance of neurotoxic dipeptide repeat proteins derived from the repeat expansion. Thus, cooperativity between gain- and loss-of-function mechanisms led to neurodegeneration. Restoring C9ORF72 levels or augmenting its function with constitutively active RAB5 or chemical modulators of RAB5 effectors rescued patient neuron survival and ameliorated neurodegenerative processes in both gain- and loss-of-function C9ORF72 mouse models. Thus, modulating vesicle trafficking was able to rescue neurodegeneration caused by the C9ORF72 repeat expansion. Coupled with rare mutations in ALS2, FIG4, CHMP2B, OPTN and SQSTM1, our results reveal mechanistic convergence on vesicle trafficking in ALS and FTD.

Acknowledgements

We thank the NINDS Biorepository at Coriell Institute for providing the following cell lines for this study: ND12133, ND03231, ND01751, ND11976, ND03719, ND00184, ND5280, ND06769, ND10689, ND12099, ND14954, ND08957, ND12100 and ND014587. We thank H. Chui and C. Miller (University of Southern California Alzheimer's Disease Research Center) and N. Shneider (Columbia University Medical Center) for control and C9ORF72 patient tissue. We thank the Choi Family Therapeutic Screening Facility for chemical screening support and the Translational Imaging Center at USC for imaging support. We thank M. Koppers, Y, Adolfs, C. van der Meer and M. Broekhoven for help with mouse breeding and kainate injection experiments. We thank S. Waguri (Fukushima Medical University) for providing the M6PR-GFP construct. We thank C, Buser for assistance with electron microscopy. We also thank S. Alworth (DRVision Technologies), K. Hebestreit and R. Bhatnagar (Verge Genomics), B. Baloh (Cedars Sinai Medical Center), J. O'Rourke (Cedars Sinai Medical Center), C. Donnelly, C. Tong, A. McMahon and Q. Liu-Michael for reagents, technical support and discussions. E.Y.S. is a Walter V. and Idun Berry Postdoctoral Fellow. K.A.S. was supported in part by a Muscular Dystrophy Association Development Grant. L.M. was supported by NIH grant T32DC009975-04. This work was supported by NIH grants AG039452, AG023084 and NS034467 to B.V.Z. R.J.P. was supported by grants from ALS Foundation Netherlands (TOTALS), Epilepsiefonds (12-08, 15-05), and VICI grant Netherlands Organization for Scientific Research (NWO). This work was also supported by NIH grants R00NS077435 and R01NS097850, US Department of Defense grant W81XWH-15-1-0187, and grants from the Donald E. and Delia B. Baxter Foundation, the Tau Consortium, the Frick Foundation for ALS Research, the Muscular Dystrophy Association, the New York Stem Cell Foundation, the USC Keck School of Medicine Regenerative Medicine Initiative, the USC Broad Innovation Award, and the Southern California Clinical and Translational Science Institute to J.K.I. J.K.I. is a New York Stem Cell Foundation-Robertson Investigator.

Author information

Author notes

Yingxiao Shi

& Shaoyu Lin

These authors contributed equally to this work.

Affiliations

Department of Stem Cell Biology and Regenerative Medicine, Keck School of Medicine, University of Southern California, Los Angeles, California, USA.

Competing interests

J.K.I. and P.A. are co-founders of Acurastem, Inc. P.A. is an employee of Icagen Corporation. J.K.I. and P.A. declare that they are bound by confidentiality agreements that prevent them from disclosing details of their financial interests in this work. S.-J.L. is a founder of DRVision Technologies and T.-Y.C. is an employee of DRVision Technologies. A.Z. and J.A.C. are co-founders of Verge Genomics and A.Z., V.H.-S., N.W. and T.G.B. are employees of Verge Genomics.